Blockade of senescence-associated microRNA-195 in aged skeletal muscle cells facilitates reprogramming to produce induced pluripotent stem cells.

Abstract

The low reprogramming efficiency in cells from elderly patients is a challenge that must be overcome. Recently, it has been reported that senescence-associated microRNA (miR)-195 targets Sirtuin 1 (SIRT1) to advance cellular senescence. Thus, we hypothesized that a blockade of miR-195 expression could improve reprogramming efficiency in old skeletal myoblasts (SkMs). We found that miR-195 expression was significantly higher in old SkMs (24 months) isolated from C57BL/6 mice as compared to young SkMs (2 months, 2.3-fold). Expression of SIRT1 and telomerase reverse transcriptase (TERT) was downregulated in old SkMs, and transduction of old SkMs with lentiviral miR-195 inhibitor significantly restored their expression. Furthermore, quantitative in situ hybridization analysis demonstrated significant telomere elongation in old SkMs transduced with anti-miR-195 (1.7-fold increase). It is important to note that blocking miR-195 expression markedly increased the reprogramming efficiency of old SkMs as compared to scramble (2.2-fold increase). Transduction of anti-miR-195 did not alter karyotype or pluripotency marker expression. Induced pluripotent stem cells (iPSCs) from old SkMs transduced with anti-miR-195 successfully formed embryoid bodies that spontaneously differentiated into three germ layers, indicating that deletion of miR-195 does not affect pluripotency in transformed SkMs. In conclusion, this study provided novel evidence that the blockade of age-induced miR-195 is a promising approach for efficient iPSC generation from aging donor subjects, which has the potential for autologous transplantation of iPSCs in elderly patients.